Ubiquitin ligases regulate protein turnover by promoting ubiquitination of substrate proteins which targets them for proteosomal degradation. CRL or Cullin-Ring ubiquitin Ligases comprise the largest class of evolutionarily conserved ubiquitin ligases. One member of this family, Cullin-3 (Cul3) plays an important role in arterial pressure regulation as mutations in the genes encoding Cul3 or Cul3 adaptor proteins cause dominant forms of hypertension. Our published evidence and preliminary data support a novel concept that vascular smooth muscle Cul3 plays a substantive role in arterial pressure regulation and its dysregulation causes hypertension. We hypothesize that Cul3 is a regulator of RhoA, a small GTPase which controls Rho kinase activity, calcium sensitivity, nitric oxide responsiveness, and contractile activity in vascular smooth muscle. Importantly, the role of vascular Cul3 expression and action in arterial pressure regulation has yet to be investigated. We will test the innovative concept that blood pressure is regulated, in part, through a Cul3-dependent pathway in vascular smooth muscle. Specifically, we hypothesize that a) impaired Cul3 activity causes hypertension, b) Cul3 mutants causing human hypertension do not properly ubiquitinate Cul3 substrates (such as RhoA), and c) Cul3 mutants act dominantly by interfering with wild-type endogenous Cul3. The aims of the project are to: 1) test the hypothesis that dominant interference of wildtype Cul3 by mutant Cul3 in vascular smooth muscle and vascular smooth muscle-specific deficiency in Cul3 results in impaired vasomotor function and hypertension, and 2) explore molecular mechanisms by which mutations in Cul3, which cause human hypertension result in impaired ubiquitination of Cul3 substrates. These studies are significant in advancing the concept that protein turnover is a regulator of vascular smooth muscle function and arterial pressure regulation, in mechanistically defining how mutations in Cul3 cause human hypertension, and in establishing mechanisms by which impaired Cul3 activity causes hypertension. The long term goal of this project is to assess if the Cul3 pathway can be used as a potential therapeutic target in hypertension.